Epstein-Barr virus (EBV) is carried by more than 95% of the adult population worldwide and is causally implicated in the development of immunoblastic lymphoma, Burkitt's lymphoma, Hodgkin's Disease, nasopharyngeal carcinoma and gastric cancer. The proximal cause of disease is the behavior of a latently infected cell. However, productive replication is critical to spread of virus within and between hosts and impacts virus load. Serologic evidence of increased virus replication is a risk factor for development of some EBV-associated tumors. The long-term goal of this research is to understand the roles that the membrane proteins of EBV play in tropism and efficiency of replication. The application focuses on understanding the multiple functions of glycoproteins gB and gNgM and the envelope protein BFRF1 and the roles they play in virus entry or spread. Glycoprotein gB has been implicated in virus assembly and is important to virus cell fusion; virus lacking gN fails to express gM and has a defect in association and possibly also in dissociation of capsids and envelope during assembly and penetration. BFRF1 interacts with BFLF2 and may play a role in exit from the nucleus. A yeast two-hybrid screen has revealed interactions between the cytoplasmic tail of gB and proteins known or thought to be involved in virus assembly. Aim 1 will test the significance of these interactions by determining if they can be reproduced biochemically, mapping the binding sites on gB and determining if mutation of binding site(s) reproduces the phenotype of a virus that lacks gB. This will be done by rescuing gB minus virus with mutated gB expressed in trans or by building mutations into a newly derived EBV-Bac. Aim 2 will test the hypothesis that gB plays a role in fusion of virions with a primary envelope and the outer nuclear membrane. gB containing mutations in the cytoplasmic tail that block virus cell fusion, or with mutations in the extracellular domain will be examined for the ability to deliver virus from the nucleus to the cytoplasm. Aim 3 will examine the phenotype of a virus that lacks BFRF1 or its partner BFLF2. Aim 4 will compare the phenotype of a virus genetically lacking gM with one that lacks gM. Yeast two-hybrid and biochemical analysis will be done to look for proteins whose interactions with the long cytoplasmic tail of gM may contribute to the phenotype of a gNgM minus virus and the phenotype of a virus that lacks the cytoplasmic tail of gM will be examined. Effects of gM on recruitment of other glycoproteins will be examined.